To promote sustainable biomass recycling and support food security, Tenebrio molitor (TM) larvae can serve as an eco-friendly source of food and feed. This study compared the survival, growth performance, and nutritional composition of TM larvae fed five diets. The control (CON) diet contained distillers’ dried grains with solubles (DDGS) and wheat bran (WB), while the experimental diets included 10–40% lignocellulose-rich organic products from rewetted peatlands (LPRP) replacing WB, with DDGS adjusted to maintain equivalent protein levels (about 21%). A total of 2500 larvae were divided into five replicates per treatment (100 larvae each). Survival exceeded 90% across all groups. Larvae fed the CON diet had a higher final body weight than those on the 30% and 40% LPRP diets (p < 0.05), with no significant differences among the CON and 10% and 20% LPRP groups. The feed conversion ratio (fresh matter) was significantly lower in the CON and 10% LPRP groups than in the other groups (p < 0.05). Larvae fed the 10% LPRP diet showed slightly higher crude protein content (55.8%) compared to the control group (54.8%) and the other treatment groups, whereas those fed the 30% LPRP diet had the highest numerical total amino acid content. Taken together, these results indicate that incorporating 10% LPRP with DDGS and WB provides the best overall balance between growth performance and nutritional quality for TM larvae, supporting sustainable production and circular economy goals.

The spread of antimicrobial resistances is a global threat. This study provides a retrospective, explorative analysis of bovine mastitis samples from 2005 to 2023, regarding the antimicrobial resistance patterns of the corresponding pathogens. 41.8

Food safety continues to be an important issue for consumer protection and public health globally. Chicken meat is considered a primary source of Salmonella and E. coli infections in humans. In recent years, phage-based biocontrol has attracted attention as a promising approach to combat these foodborne pathogens due to its advantages over traditional methods and its biological properties as a natural bactericide. Using phage-based control as a decontamination method to ensure microbial safety of food aligns with the One Health strategy for sustainable pathogen control and prevention of foodborne infections. This study aimed to develop and evaluate the effectiveness of a three-phage cocktail with optimized efficacy for simultaneously controlling Salmonella and E. coli on raw chicken filets during cold storage. To optimize the efficacy of the final phage cocktail, three phages were selected according to their host ranges and the efficiency of plating (EOP) values. They were combined in a cocktail, and the host range was expanded using the Appelmans protocol for 30 training cycles. The antibacterial efficacy of the trained three-phage cocktail was evaluated in liquid culture using a planktonic killing assay (PKA) and on raw chicken filets stored at 4 ± 0.5 °C for 72 h, employing multiplicities of infection (MOI) of 1, 10, and 100 for targeting filets contaminated with single Salmonella and E. coli strains and a mixture of both. After training according to the Appelmans protocol, the cocktail showed an expanded host range, covering 62.5% (5/8 after 30 training cycles) instead of 37.5% (3/8 before training) of the tested bacteria. The planktonic killing assay demonstrated that the trained three-phage cocktail had a significant inhibitory effect on bacterial growth of the Salmonella strains (4/4, 100%) from 3 to 6 h, while the non-trained initial three-phage cocktail’s effect was less pronounced (1/4, 25%) and lasted only 3 h. However, three of four E. coli strains (75%) were not sensitive to the three-phage cocktail after 30 cycles of the Appelmans protocol compared to two out of four strains (50%) with the non-trained initial three-phage cocktail. On raw chicken filets, significant bacterial reduction was observed when using MOI 10 and 100 of the trained three-phage cocktail. A maximum reduction of 1.56 log10 CFU/mL of Salmonella BfR 20-SA00418 and 1.48 log10 CFU/mL of E. coli 19/302/1/A after 72 h compared to placebo-treated controls were achieved using an MOI of 100. We observed a synergistic effect of the three-phage cocktail compared to single treatment, with a stronger effect on Salmonella than on E. coli strains. Using the Appelmans protocol improved the effects of the developed three-phage cocktail, leading to broader pathogen coverage. The efficacy of the developed three-phage cocktail under cold storage conditions and its ability to reduce the bacterial load in raw chicken filets highlighted its potential for extending shelf life and reducing risks for the consumer. The findings of this study demonstrate that the developed and optimized three-phage cocktail is a promising biocontrol agent for enhancing safety in raw chicken meat production.

This study describes quantitative analyses of some penicillins in goat milk after therapeutic treatment of bacterial infections. Group 1 (systemic infections, n = 5) had been treated intramuscularly with amoxicillin (AMX, 7 mg/kg b.w., 5x within 5 days), group 2 (mastitis; n = 6) intramammarily with oxacillin/ampicillin (OXA/AMP, 40/20 mg, 4x within 4 days) in one udder half. Udder half milk samples were analyzed by enzyme immunoassays, with detection limits of 1 ng/ml (AMX), 30 ng/ml (OXA), and 2 ng/ml (sum OXA/AMP). AMX residues varied between 1 and 160 ng/ml during treatment, but fell below 1 ng/ml 1 day before the end of the withdrawal period. In all samples except one, AMX concentrations were below the European Union maximum residue limit (MRL, 4 ng/ml) as early as two days after the last application. OXA levels in treated udder halves varied widely during treatment (<30 ng/ml to 1.2 mg/ml). Milk from non-treated halves also contained OXA, but concentrations were lower (<30 - 410 ng/ml). OXA fell below the MRL (30 ng/ml) 1.5 days before the end of the withdrawal period. Combined analysis of OXA and AMP showed that total residues were well below 4 ng/ml at least 1 day before the end of the withdrawal period. This is the first quantitative report on OXA excretion in goat milk after therapeutic application under real life conditions of goat milk production. The milk of all analysed goats complied with MRL regulations after the withdrawal period of 4 days (OXA) and 4.5 days (AMX).

Diarrheal disease caused by Bacillus cereus is driven by pore-forming enterotoxins, with hemolysin BL (Hbl) and non-hemolytic enterotoxin (Nhe) as major virulence factors. Toxin structure and mode of action are well described, while especially human intestinal epithelial responses remain incompletely characterized. With the co-culture system of Caco-2 and mucus-producing HT29-MTX cells (9:1), this study describes a suitable model for investigating enterotoxin effects. Cell line-, dose- and enterotoxin-dependent changes in cell viability were detected. Undifferentiated Caco-2 generally showed higher susceptibility than HT29-MTX cells. Twenty-four-hour co-cultures were more susceptible than 28-day co-cultures but displayed partial recovery after toxin removal. Enterotoxin exposure caused a time-dependent decline in transepithelial electrical resistance, consistent with barrier dysfunction. In Ussing chamber experiments, enterotoxin-containing supernatants, especially Nhe+Hbl, increased short-circuit currents, indicating altered electrogenic ion transport consistent with pro-secretory phenotypes relevant to diarrhea. Transcriptome analyses demonstrated a pronounced time- and dose-dependent response toward Nhe+Hbl. Two-hour sublethal enterotoxin exposure elicited few transcriptional changes, whereas lethal enterotoxin exposure rapidly suppressed proliferation- and cell-cycle-associated programs. Pathway analyses consistently highlighted TNFα/NF-κB-driven inflammation, MAPK-associated stress signaling, and p53/apoptosis pathways; metabolic remodeling and senescence-/autophagy-related processes became more apparent at longer exposure (8 h) and/or higher toxin levels. Dose-dependent caspase-3/7 activation, LDH release, and minimal caspase-1 activation were also consistent with early inflammatory response, apoptotic cell death and membrane damage. Collectively, this study integrates barrier function, ion transport, cell viability, and transcriptomics to define host intestine epithelial response programs triggered by B. cereus enterotoxins and to connect toxin exposure to barrier dysfunction and diarrheal pathophysiology.

Trueperella (T.) bernardiae has been only rarely identified from animals, and its genomic diversity remains poorly characterised. This study analysed two canine-derived isolates initially suspected of representing T. bernardiae using phenotypic, molecular and whole-genome-based approaches. While the phenotypic profiles showed overall similarity to the type strain, several characteristics differed. The control strain was reliably identified phenotypically, whereas two isolates could not be identified beyond the genus level. Additionally, despite their high 16S rRNA gene similarity to the type strain, phylogenomic analyses consistently placed both isolates in a separate lineage. In the whole-genome sequencing (WGS)-based phylogeny, the two isolates formed a distinct cluster closest to T. bernardiae DSM 9152ᵀ and T. bernardiae UMB8254. Whole-genome comparisons supported this distinction, with average nucleotide identity values (ANIb and ANIm) of 93.98-94.69% and digital DNA-DNA hybridization (dDDH) values of 57.2-57.6% relative to T. bernardiae DSM 9152ᵀ. These findings indicate that the two isolates show genomic divergence from known Trueperella species, raising the possibility that they form a separate lineage. This study also provides the first whole-genome comparison of human- and animal-derived T. bernardiae isolates, expanding the limited genomic framework for this understudied genus.

Enzyme immunoassays (EIAs) for small molecules often suffer from limited sensitivity or high non-specific background. This study presents a reproducible strategy to improve EIA performance by optimizing hapten-protein conjugation conditions. Using an immunoassay for the mycotoxin α-cyclopiazonic acid (α-CPA) as a case study, key parameters of the formaldehyde-mediated (Mannich) condensation reaction for synthesizing a hapten-bovine serum albumin conjugate were systematically investigated. Variation of reaction temperature (8-37°C) and time (0.25-72 h) showed that short reaction times at low temperature markedly improved assay performance. Under optimized conditions (8°C, 15 min), the detection limit improved from >100 ng/mL to 2.4 ± 0.2 ng/mL, with a strong reduction in non-specific background. The results demonstrate that targeted optimization of hapten-protein conjugation can significantly enhance EIA sensitivity and specificity, which helps to reduce the number of laboratory animals for antibody production. The approach is readily transferable to other indole-containing mycotoxins and was successfully applied to determine α-CPA in fungal agar plugs from mold-ripened cheeses. This study highlights an important aspect of conjugation conditions during immunoassay optimization. •Systematic optimization of formaldehyde-mediated hapten-protein conjugation for EIAs •Improved assay sensitivity and background without new antibody production •Transferable strategy demonstrated for α-CPA analysis in complex fungal samples.

AIMS:Zoonotic infections with Salmonella spp. transmitted from reptiles to humans are an increasing concern due to the growing number of documented cases and the close contact between humans and reptiles. Reptiles, such as bearded dragons (Pogona vitticeps), frequently carry Salmonella enterica asymptomatically as part of their intestinal microbiota.Given the rise of antibiotic-resistant Salmonella strains in reptiles, bacteriophages (phages) may provide a targeted and sustainable alternative for preventing reptile-to-human transmission. METHODS AND RESULTS:Seventeen phages were isolated from ten of eighteen fecal samples collected from bearded dragons. Seven of these phages were selected for further analyses. Host range assays on 41 S. enterica and nine non-Salmonella isolates revealed a narrow spectrum: phages infected up to 63.4% of Salmonella isolates and lysed one non-Salmonella strain. Planktonic killing assays at 25 and 37°C showed pronounced bacterial growth reduction, with 8 of 12 significant inhibitions observed at 37°C. Phage cocktails generally showed stronger inhibition than individual phages. Electron microscopy and whole-genome sequencing identified six Myovirus-like and one Siphovirus-like phage, all temperate with integrase or transposase genes despite lytic activity. CONCLUSIONS:This study expands knowledge of S. enterica-specific phages from reptiles, detailing host specificity, morphology, and genomic features. While in vitro results are promising, in vivo efficacy may be influenced by host physiology, immunity, and microbiome interactions. The predominance of temperate phages may limit direct therapeutic use, though low lysogeny rates in related phages and genetic engineering advances may enable future applications.

The genus Arcanobacterium has expanded rapidly, with an increasing number of species described from domestic and wildlife hosts, underscoring the need for clearer delineation of species boundaries. In this study, 12 isolates initially presumed to belong to Arcanobacterium canis were examined using comparative phenotypic and genomic approaches to clarify their taxonomic position. The isolates showed partial phenotypic similarity to the type strain A. canis DSM 25104ᵀ, including consistent haemolysis, but differed in several enzymatic reactions and carbohydrate utilization patterns. Matrix-assisted laser desorption/ionization time-of-flight MS identified all isolates at the genus level, with only one displaying spectral similarity to A. canis. Phylogenetic analysis of five gene targets revealed that although the isolates shared sequence similarity with the type strain A. canis DSM 25104ᵀ, they consistently formed a separate clade. Core-genome phylogeny likewise placed the isolates on an independent branch. Pangenome analysis identified 2,178 gene clusters (58.8% core genes) and demonstrated that the cytolysin-like gene present in the type strain A. canis DSM 25104ᵀ is completely absent from all 12 isolates. Digital DNA-DNA hybridization and average nucleotide identity values fell below accepted species-level thresholds, indicating that the isolates represent a genomically distinct lineage within Arcanobacterium. Because chemotaxonomic analyses required for formal species description were not performed, the taxonomic status of this lineage remains unresolved. Nevertheless, the present findings refine species boundaries within Arcanobacterium and provide a robust foundation for future taxonomic and comparative genomic studies.

The lactic acid bacterium Leuconostoc carnosum is a noteworthy spoilage organism in the context of the food industry, given its prevalence in meat products. Furthermore, there is also evidence that L. carnosum may have a key role in salt-and nitrite-reduced product formulations. At present, there do not exist any selective culture methods for L. carnosum, and only a limited number of publications are available on molecular diagnostic detection methods. Therefore, in this study a realtime PCR assay was developed and validated for the purpose of tracking the exact growth behaviour at species level of L. carnosum in meat products. The quantitative polymerase chain reaction (qPCR) method based on the lysA gene showed a specificity of 100% when tested with bacterial strains of the target species and relevant non-target species. The detection limit in artificially contaminated cooked ham samples was 6 x 102 cfu/g. The newly qPCR was validated in a native sample context by determining the quantitative growth of the target species using the new qPCR and a literature-based reference qPCR. The findings show that the two methods produce similar results in terms of detection and quantification, demonstrating a close agreement between them. This study successfully implemented a TaqMan qPCR method for the detection of L. carnosum in cooked ham and is valuable for facilitating the execution of challenge tests concerning the spoilage process of salt-and nitrite-reduced meat products. This will substantially support further research in the development of effective strategies to ensure food quality and safety.

Delivered ready-to-eat foods have grown increasingly popular in many countries worldwide. Takeaway meals are subjected to transport and storage under variable conditions. This study examined the influence of packaging type (open, semi-open and sealed) and storage conditions (6 °C, 20 °C and 30 °C for 30 min, 2 h and 24 h) on the microbiological safety and quality of grilled pork, with particular focus on cross-contamination from contaminated meat, packaging or side dishes. The total bacterial count of non-inoculated grilled pork continuously remained below the detection limit (0.7 log CFU/g), confirming effective thermal processing before inoculation. Either meat or the packaging surface was inoculated with Escherichia coli, Brochothrix thermosphacta or Yersinia enterocolitica. Furthermore, pasta inoculated with Bacillus cereus was combined with non-inoculated meat. Bacterial growth was significantly influenced by storage temperature, duration and packaging type. Storage at 20 °C and 30 °C, particularly for 24 h, resulted in markedly higher bacterial counts than storage at 6 °C. Semi-open and sealed packaging generally promoted greater bacterial growth and cereulide production than open systems. During storage, cross-contamination of packaging or pasta onto meat was detected. In conclusion, short storage times combined with refrigeration and non-sealed packaging effectively minimized microbial growth and toxin formation. These findings underscore the importance of proper packaging selection and short-term, chilled storage of ready-to-eat foods. This is essential not only to control multiplication of pathogens and spoilage microorganisms, but also to prevent production of stable toxins that cannot be eliminated by subsequent reheating of the food.

The COVID-19 pandemic has led to a significant increase in food delivery services due to the heightened demand for ready-to-eat meals. This study investigated the effects of different packaging systems and storage conditions on the microbiological, chemical, and sensory properties of turkey meat. Three packaging systems (open, semi-open, and sealed) were evaluated, and the samples were stored at three temperatures (6 °C, 20 °C, and 30 °C) for various durations (30 min, 2 h, and 24 h). In addition to the investigation of noninoculated samples, separate experiments were conducted in which either the turkey meat or the inner surface of the packaging was inoculated to assess microbial behavior and the potential for cross-contamination. Noninoculated samples remained below the detection limit across all conditions, demonstrating the effectiveness of thermal processing at 75 °C. Inoculation experiments with Escherichia (E.) coli, Brochothrix (B.) thermosphacta, and Campylobacter (C.) jejuni revealed that packaging systems with minimal or no air exchange (semi-open, sealed packaging) promoted bacterial growth at higher temperatures (20 °C, 30 °C). After 24 h at 30 °C, E. coli and B. thermosphacta reached 6.0 - 8.5 log CFU/cm2 in sealed packaging, while C. jejuni showed minimal growth (≤2.8 log CFU/cm2) due to its microaerophilic requirements. Open packaging limited bacterial growth, particularly at lower temperatures (6 °C). Sensory evaluation also showed significant differences regarding storage conditions. The study demonstrates that a short storage time in combination with low temperatures and packaging that allows for air exchange reduces the risk of foodborne illness most effectively. In contrast, packaging that allows minimal or no air exchange with the environment appears to be more favorable in terms of sensory attributes. These results highlighted the need to balance microbiological safety and sensory quality when selecting packaging systems for takeout meat products.

Mastitis remains the most frequent disease in dairy cows, affecting both animal welfare and the profitability of dairy farms. In the present investigation, we retrospectively analysed 102,179 cow's milk samples, which were object of routine diagnostics at the Department of Milk Hygiene, Institute for Food Quality and Food Safety, University of Veterinary Medicine Hannover, from 2005 to 2023, regarding the occurrence of major mastitis pathogens. On average, approx. 48% of all tested samples were bacteriologically positive. Differences in the occurrence of positive findings were described regarding the season, the submitter, the reason for submission, as well as the udder quarters. Generally, the 13 most common groups of pathogens accounted for 94.0% of all positive samples. Non-aureus staphylococci and mammaliicocci (23.6%) were detected most frequently, followed by Streptococcus uberis (14.8%), Escherichia coli (13.2%), Corynebacterium spp. (9.5%), Staphylococcus aureus (6.7%), yeast (6.1%), and Enterococcus spp. (5.6%). While a slight decline was recorded for most of these pathogen groups, S. uberis and E. coli showed a significant upward trend throughout the investigation period. Moreover, a constantly high percentage of samples contaminated with more than three different colony forms was found. Monitoring long-term development of the mastitis pathogen spectrum as well as its antibiotic resistance patterns is of utmost importance to enable adaptation of hygiene and treatment strategies. This includes appropriate training of the treating veterinarians with regard to sterile milk sampling.

The use of sodium chloride (NaCl) in the food industry is increasingly being criticized as it is suspected of having potentially harmful properties. For this reason, various methods are being pursued to research alternatives to its use. However, the reduction of NaCl and nitrite harbors microbiological risks and can also have an impact on the technological properties of the products. In the experiment described here, various microbiological and physico-chemical parameters of raw sausages with different NaCl and nitrite contents were examined over a storage period of 140 days. The aim was to investigate the shelf life of raw sausages produced with different recipes in comparison to a raw sausage with conventional NaCl and nitrite contents (reference) in order to be able to make a statement about the possibilities and limits of the reduction of NaCl and nitrite.

In the interest of enhancing consumer health and limiting the risks associated with excessive salt consumption and nitrosamine intake, there is an ongoing initiative to reduce the use of salt and nitrite in meat products. Since important microbial barriers are compromised in their capacity to function effectively, NaCl- and nitrite-reduced products also run the risk of increasing the growth of potentially pathogenic microorganisms, such as Clostridium botulinum. In consideration of the necessity to investigate the growth behaviour of the pathogen in sodium chloride (NaCl) - and nitrite-reduced meat products in future challenge tests, a real-time PCR assay was developed and validated for the qualitative and quantitative detection of the C. botulinum surrogate Clostridium sporogenes. The quantitative polymerase chain reaction (qPCR) method based on the pelF gene showed a specifity of 100% when tested with bacterial strains of the target species and relevant non-target species. The detection limits in artificially contaminated raw sausages were 8.6 cfu per reaction and 430 cfu/g. The validation of the newly established TaqMan qPCR assay was conducted while investigating the impact of the ripening process and long-term storage of raw sausage with different salt and nitrite contents on the growth of C. sporogenes. The qPCR results were then compared to those of classic cultivation, serving as the reference method for validation. In summary the results of the qPCR showed a good correlation with those of the cultural method for the simulated conditions. However cell counts after ripening were slightly higher, when measured using qPCR. This study presents the successful implementation of a qPCR method for the detection of C. sporogenes in raw sausage and offers a significant opportunity for further research that is required in order to determine the possibilities of reducing salt and nitrite in meat products.

Cow’s milk contains A1- and A2-β-caseins. The breakdown of A1-β-casein produces β-casomorphin-7 (BCM-7), a peptide with opioid-like properties that is associated with health aspects. In addition, A1- and A2-β-casein have different technological properties. The aim of the present study was to investigate whether cheese produced from the milk of homozygous A1A1 and A2A2 cows varies in terms of its physicochemical parameters and BCM-7 concentration. These parameters were analyzed during initial cheese processing, six weeks of ripening and 84 days of storage, including additional microbiological analyses during the storage period. The pH values of the A1A1 cheeses were higher than those of the A2A2 cheeses from the beginning of production until the starter culture bacteria were added. The yellowness values of the A1A1 cheeses were lower until the salt bath treatment. Water activity, lightness, hardness, fat, protein, NaCl and dry matter content, as well as color and microbiological parameters, were not affected by the β-casein genotype. BCM-7 concentrations were higher in the A1A1 cheeses after pressing and during ripening. We found mainly comparable quality characteristics and slightly different BCM-7 levels in the A1A1 and A2A2 cheeses. From this point of view, both varieties are equally suitable for cheese production.

The brilliant black reduction test (BRT) is a first and rapid screening test routinely used across the German dairy industry for the detection of antimicrobial substances, especially antibiotics, in cow's milk samples. This study aimed to determine if the three test systems BRT Inhibitor Test, BRT MRL-Screening Test and BRT hi- sense can also be applied to buffalo and horse milk. For buffalo milk, the detection limits of all antibiotics under investigation were at or below the European maximum residue levels (MRL). Previous freezing of the antibiotics in buffalo milk showed comparable results. While the BRT hi-sense is not recommended for horse milk, detection limits at or below the MRL were reached with the BRT Inhibitor Test and/or the BRT MRL-Screening Test for penicillins, neomycin, gentamicin, as well as streptomycin (BRT MRL-Screening Test only). In general, it must be noted that the incubation times of the test systems need extending compared to cow's milk and that horse milk must be heated before application.

BACKGROUND:Antimicrobial stewardship has become vital given the progressive emergence of multidrug-resistant bacteria, and novel approaches to the treatment of bacterial infections are needed. Recently, reported synergistic effects of antibacterial drugs and bacteriophage therapy have revealed promising applications for the management of meticillin-resistant staphylococcal infections. OBJECTIVES:The objective of this study was to investigate the response of meticillin-resistant Staphylococcus pseudintermedius (MRSP) to treatment with a newly isolated, lytic MRSP-specific bacteriophage. Furthermore, a postulated synergism between phage and fusidic acid was examined in a canine ex vivo dermis model. MATERIAL AND METHODS:Skin was harvested from the lateral thorax of a euthanised dog, clipped, the subcutis removed, and epidermis cleaved via a modified salt-split technique. The ex vivo dermis model established in Franz diffusion cells was inoculated with 1 × 107 colony-forming units (cfu) of a clinical MRSP strain for 16 h. Then, experimental groups were treated with phage vB_SpsS_LmqsKl44-4 at a concentration of 2 × 106 plaque-forming units and fusidic acid 0.4 mg alone or in combination for an additional 8 h. RESULTS:Histopathological results showed that colonies of MRSP reached the superficial dermis and entered hair follicles. Co-treatment with fusidic acid and phage significantly reduced the amount of MRSP after 8 h. CONCLUSIONS AND CLINICAL RELEVANCE:In conclusion, topical co-treatment with fusidic acid and a phage could be a promising approach to the treatment of canine MRSP pyoderma.

Actinomyces weissii was first isolated from the oral cavity of dogs in 2012. This study characterised the phenotypic and genotypic features of 11 additional strains obtained from diverse pathological lesions in companion animals. A multi-faceted approach was employed, combining culture techniques, biochemical profiling, MALDI-TOF MS, real-time PCR, sequencing of genetic markers, and whole-genome analysis. Despite minor phenotypic variation, MALDI-TOF MS and genotypic analyses consistently confirmed species identity and revealed intraspecies diversity. Whole-genome sequencing revealed four haemolysin family protein genes, and all isolates exhibited complete β-haemolysis. Pan-genome analysis defined a conserved core genome of 1559 genes, shared by all isolates and the reference strain, including haemolysin-related genes. Phylogenetic comparisons placed A. urogenitalis and A. trachealis as closest relatives. These findings broaden the host spectrum of A. weissii, with the first isolation from a cat, suggesting that this species may also occur in felines and merits consideration in veterinary diagnostics. Its presence in companion animals raises the possibility of zoonotic risk, and together with the detection of haemolysin family protein genes, underscores the need for further investigation to clarify its pathogenic significance and potential impact on veterinary practice and public health.